The ability to convert proinsulin to insulin using trypsin and carboxypeptidase B has been recognized for several years [see, e.g., Kemmler, W., Clark, J. L., Borg, J. and Steiner, D. F., Fed. Proc. 30 (1971) 1210; Kemmler, W., Peterson, J. D., and Steiner, D. F., J. Biol. Chem., 246 (1971) 6786-6791]. An ongoing difficulty with this conversion methodology has been and continues to be the presence of substantially large amounts of difficultly-removable by-products in the reaction mixture. Specifically, in the conversion of human pro-insulin to human insulin, a large amount (about 4-6%) of Des-Thr(B30) human insulin is formed. This by-product, differing from human insulin by the absence of a single terminal amino acid, is, if capable of being separated from the product mixture at all, separated only by difficult and cumbersome methodology.
With the advent of recombinant DNA technology, for the first time large amounts of human proinsulin have become a reality. In using the human proinsulin as an intermediate in the production of insulin, a solution to the Des-Thr(B30)-hI impurity problem has become mandatory. Either one could seek ways to achieve purification of the human insulin from the contaminating Des-Thr(B30)-hI or seek a conversion process, the conditions of which minimize formation of the latter. It is to a new process for converting a human insulin precursor to human insulin with minimal formation of Des-Thr(B30)-hI that the present invention is directed.